The capability to detect and diagnose infections by non-human primate viruses is far less than optimal and in cases non-existent. As Specific Pathogen Free monkey colonies are developed, it will be essential that maximally sensitive tests for monkey viruses are available to develop and monitor such colonies. The overall goal of the proposed research is to develop basic molecular data for BV of macaques and to utilize these results to develop diagnostic tests that are more sensitive, more specific and safer than those currently available. To achieve these goals, the following experiments will be performed: 1) An indicator cell line will be constructed for diagnostic use in virus isolation which expresses GFP when infected by BV, making virus isolation testing for BV a more sensitive and safer procedure. 2) A lethal-deletion mutant strain of BV (ABV) will be constructed by deleting an essential structural gene and constructing a complementing cell line in which ABV can replicate. This system (ABV + complementing cell line) will make preparation of BV antigens and DNA much safer. 3) Major BV immunogens will be identified by assessing the macaque immune response to individual BV proteins. This information will allow development of effective vaccines and recombinant antigens for use in diagnostic tests. 4) Specific glycoprotein genes of BV will be expressed using an inducible mammalian expression system, and the expressed proteins used to develop immunciassays. 5) A TaqMan assay will be developed that can detect and differentiate herpes simplex virus-1 (HSV I), herpes simplex virus-2 (HSV2) and BV. This will provide an extremely sensitive means of diagnosing human BV cases. Successful completion of the proposed experiments will provide a number of reagents that will allow much safer detection of infectious BV in clinical samples, safer preparation of BV antigen for diagnostic tests, more sensitive and specific serological tests for detection of anti-BV antibodies in both macaque and HSV-positive human sera, and basic information regarding the antigenic determinants of BV which may induce protective immunity against infection.